Detection of clinically relevant antibiotic-resistant bacteria in shared fomites, waste water and municipal solid wastes disposed near residential areas of a Nigerian city

Studies investigating environmental hotspots of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in Nigeria are limited. This study was designed to assess various environmental sources and commonly touched surfaces as potential carriers of ARB and ARGs with implications for public health. A total of 392 samples, including sewage (36), sludge (36), diapers (20), plastics (20), water sachet polythene bags (20), food wastes (20), soil beneath dump sites (20), and frequently touched surfaces such as restroom floors (80), corridors (24), door handles (56), and room floors and walls (60), were collected and screened for the presence of resistant bacteria carrying genes such as bla KPC , bla NDM-1 , bla CMY-2 , bla IMP , bla OXA66 and MecA. Additionally, we employed standard techniques to detect methicillin-resistant Staphylococcus aureus (MRSA) and extended-spectrum β-lactamase (ESBL)-producing Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii . We also evaluated the effectiveness of routine disinfection procedures in eliminating ARB from restroom floors. Our findings revealed that sewage, sludge, diapers, food wastes and restroom floors are frequently contaminated with highly and moderately resistant strains of E. coli, K. pneumoniae, P. aeruginosa and MRSA. Notably, we identified two variants of the bla OXA51-like gene (bla OXA-66 and bla OXA-180 ) in A. baumannii isolated from these environmental sources. Furthermore, we detected seven ESBL- K. pneumoniae , five ESBL- A. baumannii , two ESBL- E. coli and one ESBL- P. aeruginosa , all carrying one or more ARGs (bla KPC , bla NDM-1 , bla CMY-2 ), in isolates recovered from sewage, sludge, restroom floors and plastics. It is of note that ARB persisted on restroom floors even after disinfection procedures. In conclusion, this study highlights that environmental wastes indiscriminately discarded in residential areas and shared surfaces among individuals are heavily colonized by ARB carrying ARGs of significant public health importance.


INTRODUCTION
The open dumping of municipal solid waste (MSW), without segregation and treatment, is a significant concern due to its hazardous contents of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs), and other antibiotic resistancedriving agents (ARDAs).Increased interest in determining the role of the environment in driving and spreading ARB and ARGs has led to a renewed interest in surveying different environmental wastes and frequently touched fomites for carrying ARB and ARGs of public health concern.
The overuse of antibiotics in humans and animals has led to continual release of antibiotics through excretory products into terrestrial (e.g.dumpsites) and aquatic (e.g.stagnated waters) ecosystems, which has resulted in the evolution of new ARB and ARGs of great concern to public health [1,2].As MSWs are not properly disposed of in most Nigerian cities, the proximity of illegal disposal sites to residential areas makes it easy for microbial populations that have accumulated in the wastes to crosscontaminate humans, animals and shared fomites.In addition to MSW, wastewater and its derivatives (sewage from sewers and sludge from gutters) are also discharged into the environment from sources, such as laundries, kitchens, agricultural land (in the form of runoff) and industries (in the form of effluents) and are often contaminated with different bacteria, and unquantifiable amounts of ARDAs such as residual antibiotics, disinfectants, metals and other toxic chemicals [3,4].
In Nigeria, while sewage and sludge frequently stagnate in drainage systems in the form of gutters, MSWs are regularly made up of plastics, water sachet polythene bags, diapers, food wastes, metals, etc., and end up in open areas and drainages.
The presence of high densities of active ARB and ARGs in MSW and fomites could serve as hotspots from where additional ARB could evolve through horizontal transfer of ARGs to other bacteria or place humans and animals scavenging or living in surrounding areas at risk of acquiring and disseminating ARB and ARGs through direct contact.[5,6] Despite the worldwide distribution and endemicity of methicillin-resistant Staphylococcus aureus (MRSA) and extended spectrum beta-lactamase (ESBL)-producing Escherichia coli (ESBL-E.coli), Klebsiella pneumoniae (ESBL-K.pneumoniae), Pseudomonas aerugiosa (ESBL-P.aeruginosa) and Acinetobacter baumannii (ESBL-A.baumannii) in clinical settings [7][8][9], little is known regarding the level of colonization of the major components of MSW, waste water derivatives (WWDs) and fomites with ARB in Nigeria and whether they also harbour ARGs of clinical importance.Previous isolation of carbapenem-resistant Enterobacteriaceae in rural communities with no prior use of carbapenem in Nigeria may suggest an environmental carriage and reservoirs which can be transferred to hospital environments via sick or asymptomatic carriers.[9] A better understanding of burden, ecological niches and reservoirs of MRSA and ESBL-producing bacteria in indiscriminately discharged environmental wastes and shared fomites would help in managing and controlling their spread in hospitals and communities.The objectives of the present study were: (i) to screen major components of MSW, WWDs and shared fomites in a Nigerian city for ARB carriage; (ii) to screen the ARB isolated for carriage of some ARGs of public health concern; and (iii) to evaluate the efficacy of restroom floor disinfection procedures routinely used in the university in reducing the burden of bacterial contaminants.

Study setting and sample collection
Prior to the start of the study, an ethical clearance was sought and provided by the ethical committee of Kano State Ministry of Health.The study was conducted in the Department of Microbiology, Bayero University Kano (BUK), Center for Biotechnology Research (CBR), BUK and Microbiology Department of Aminu Kano Teaching Hospital Kano-Nigeria.A total of 392 samples/ swabs were collected between March and June 2021 as follows.

Solid waste and waste water sampling
Samples of plastics, diapers, sachet water polythene bags, food wastes and soil from different MSW illegal disposal sites situated around residential areas were collected using sterile forceps and a spatula at random and placed into sterile glass plastic containers.Meanwhile, samples of sewage and sludge were also collected from gutters and open sewers around residential areas at Dorayi Karama, Hotoro Tsamiyar Boka and hostels for men of Bayero University Kano, Nigeria.Sites of sampling and the nature of discharged wastes are shown in Fig. 1.Samples were placed into sterile containers and were transported immediately to the laboratory for processing.In the laboratory, three different surfaces of plastics, diapers and sachet water polythene bags were swabbed to form a composite sampling using a sterile cotton-tipped swab stick.

Fomite sampling
A 4 day longitudinal sampling of fomites in Nigerian University hostel was undertaken.Samples were collected from two categories of surfaces.
(1) Restroom floor: samples were collected across two blocks of hostels comprising five restrooms per block.Restroom floors were sampled with moist swabs from two locations (floors around the toilet seat and floors around the door) in the morning (8 a.m.) before daily cleaning and 1 h after routine cleaning.In addition, samples from the floor of a corridor between the restroom and rooms were also collected.
(2) Shared rooms: walls, floors and door handles of rooms shared by students were also sampled using similar techniques.

Isolation, identification and screening of MRSA and ESBL-producing bacteria
Sewage and sewage samples were appropriately diluted before inoculation onto nutrient agar plates, which were subsequently incubated at 37 °C for 24 h.Following incubation, suspected isolates of S. aureus, K. pneumoniae, P. aeruginosa and E. coli were sub-cultured onto mannitol salt agar, EMB and MacConkey agar plates.
For each isolate, observations were made regarding its colonial characteristics, including size, shape, colour, margin, elevation and opacity.Additionally, morphological characteristics such as Gram reaction, cell shape and arrangement were determined according to established methods.The identity of the isolates was further confirmed using the API 20E kit (bioMérieux).
Confirmation of A. baumannii relied on the detection of bla OXA-51-like genes in genomic DNA obtained through the boiling method.Antibiotic susceptibility patterns of all the isolates were determined following the standards specified by the Clinical Laboratory Standards Institute (CLSI).In brief, approximately 150 µl of standardized nutrient broth for each bacterium was streaked onto Mueller-Hinton agar plates (Oxoid) in duplicate.Sixteen antibiotic discs (all from Oxoid) were placed on each plate, including amoxicillin, ampicillin, ceftriaxone, cefoxitin, cefotaxime, cefepime, imipenem, ciprofloxacin, levofloxacin, gentamicin, kanamycin, tetracycline, tigecycline, clindamycin, erythromycin and colistin.After 24 h of incubation at 37 °C under ambient conditions, the diameters of inhibition zones were measured and interpreted according to species-specific CLSI 2018 breakpoints [10].Furthermore, S. aureus isolates were screened for MRSA phenotypically using cefoxitin susceptibility and genotypically by amplification of mecA genes.Resistant isolates of E. coli, P. aeruginosa, K. pneumoniae and A. baumannii were screened for ESBL production using the double disc synergy test, as described in CLSI 2018 [11].

Detection of ARGs
The ARGs considered in this study include bla KPC , bla NDM , bla IMP and bla CMY-2 , which confer resistance to commonly used β-lactam and carbapenem antibiotics in the study area.Plasmids were extracted from resistant E. coli, K. pneumoniae, P. aeruginosa and A. baumannii using a commercial kit (Zymo Research).Subsequently, the selected ARGs were amplified using conventional PCR with their respective oligonucleotides (Table 1).For plasmid DNA extracted from E. coli, K. pneumoniae and P. aeruginosa, PCR amplification targeted fragments of bla NDM , bla IMP , bla KPC and bla CMY-2 , measuring 476, 587, 882 and 106 bp, respectively.Each PCR consisted of 2 µl of template DNA, 1 µl of primers (at final concentrations of 0.66 µM each) and 12.5 µl of X2 Dream Taq Master Mix (Thermo Scientific), in a final volume of 25 µl.The PCRs underwent 30 cycles using previously described cycling conditions [12].PCR amplicons were visualized on a 1.5 % agarose gel stained with ethidium bromide for 42 min (150 V, 400 A), and the gel image was captured using a documentation device (OmniDOC Gel Documentation System; Cleaver Scientific).Samples displaying a band were selected, purified using a DNA and PCR purification kit (GDSBios) and subsequently sequenced.
For sequencing, 5 µl of purified amplicon was subjected to Sanger sequencing on a 3730xl DNA Analyzer (ThermoFisher Scientific) at Inqaba Biotech.Sequence chromatograms were visually inspected for quality using FinchTv (version 1.4).These sequences were then compared to existing sequences in GenBank (https://www.ncbi.nlm.nih.gov/genbank/) using blast with default parameters to search in the 'nucleotide collection (nr/nt)' database (GenBank) for highly similar sequences (megablast) (TBLASTX, RRID:SCR_011823).To determine the specific bla OXA-51-like gene variant of the A. baumannii isolates, the coding region of the gene was amplified with bla OXA69 primers and fully sequenced according to method described by Pfeifer et al. [12] and sequence assembly was performed de novo using the Geneious bioinformatic tool for contig assembly.The consensus sequence was extracted, copied and blasted using the translated nucleotides to protein tool (Blastx).

Efficacy of restroom floor disinfection process in reducing ARB
Because the restroom floors are often disinfected every day, the efficacy of the type of disinfectants used in removing or reducing bacteria or ARB were tested by checking for the presence of ARB and other bacteria 1 h after the cleaning process [23].

Data analysis
The results are presented in frequency distribution tables and figures.Statistical analysis was performed using the SPSS (RRID:SCR_002865) statistical package (SPSS).A chi-square test was used to compare differences between groups.Statistical significance was set at P<0.05.

Isolation of bacteria from environmental wastes and shared fomites
Out of the total 392 samples that were screened, 191 (48.7 %) produced positive cultures of S. aureus, E. coli, P. aeruginosa, K. pneumoniae and A. baumannii, each at varying rates.The counts of bacteria for each sample type are given in Table 2.Among all the sampled areas, including MSW, WWD and fomites, the highest bacterial recovery rates were observed in sewage (88.9 %), the sand where samples were disposed of (85 %), sludge (75 %) and restroom floors (37 %).Furthermore, A. baumannii was more frequently recovered in WWD, with rates of 11.1 % in sewage and 8.3 % in sludge, as well as in fomites, with rates of 12 % on door handles and 10 % on restroom floors, compared to MSW, where it was only isolated in the sand sample (5 %).

Antibacterial susceptibility of isolated bacteria to antibiotics
Antibiotic sensitivity testing conducted on 39 E. coli, 55 K. pneumoniae, 52 S. aureus, 16 P. aeruginosa and 20 A. baumannii samples revealed that they exhibited high suscebtibiity (90-100 %) to cephalosporins and the carbapenem (imipenem).In contrast, the majority of these isolates demonstrated resistance to ampicillin and amoxicillin, and showed intermediate resistance to fluoroquinolones.Notably, all A. baumannii strains exhibited complete resistance to penicillins (ampicillin and amoxicillin), tetracycline, tigecycline, erythromycin, gentamicin, kanamycin and chloramphenicol, but remained susceptible to colistin and imipenem.

ESBL and MRSA detections
All samples were assessed for the presence of Gram-negative bacteria displaying resistance to at least penicillin, cephalosporin and/or fluoroquinolone antibiotics, with a focus on screening for ESBL production.ESBL production was confirmed in only seven K. pneumoniae isolates obtained from diapers, food wastes and restroom floors, five A. baumannii isolates from sludge and restroom floors, two E. coli isolates from sewage and one imipenem-resistant P. aeruginosa isolate also from sewage.Additionally, 10 of the isolated S. aureus, originating from restroom floors, door handles, polythene bags and sewage, were identified as MRSA, as detailed in Table 4.

ARG detection in selected bacteria
Two ARGs, namely bla CMY-2 and bla NDM were found within bacteria that were isolated from various sources, including sewage, sludge, plastics and restroom floors.Among these, bacteria isolated from sewage and restroom floors exhibited a higher prevalence of ARGs compared to other samples.Specifically, three E. coli strains isolated from sewage, sludge and restroom floors harboured the bla CMY-2 , bla IMP and bla NDM-1 genes, respectively.Notably, one ESBL-K.pneumoniae isolated from sewage co-harboured the bla KPC  and bla NDM-5 genes on its plasmid.ESBL-P.aeruginosa and non-ESBL producers carried the bla KPC and bla CMY-2 genes, respectively (Table 5).The sequences of bla NDM from two isolates showed a 99.82 % identity to the known NDM-5 gene sequence, whereas the sequence from the E. coli isolated from the restroom floor revealed the bla NDM-1 gene.The bla CMY-2 and bla KPC genes exhibited nucleotide sequence identities ranging from 98 to 100 % with related strains in the database.Additionally, two A. baumannii strains isolated from sludge and a door handle harboured the bla KPC and bla CMY-2 genes, respectively.When the assembled bla OXA-51-like gene sequences from A. baumannii were subjected to Blastx database searchs, it was found that the isolates encode protein variants of bla OXA-66 , known to be associated with clinical international clone A. baumannii lineages, whereas the isolate from the door handle belonged to the bla OXA-180 variant.Finally, six out of the 10 MRSA strains were found to carry the mecA genes.

Efficacy of floor disinfection process routinely used in reducing burden of selected bacteria
A comparison was made between before and after a routine disinfection process of restroom floors, during which an unknown amount of disinfectant was used, to assess its effectiveness in removing ARB from the restroom floors.Nearly all the samples collected from the restroom floors in the morning before cleaning showed positive cultures for S. aureus, E. coli, K. pneumoniae, P. aeruginosa and A. baumannii throughout the study period.The floors exhibited higher contamination levels of S. aureus, E. coli and K. pneumoniae compared to P. aeruginosa.Specifically, K. pneumoniae and S. aureus were significantly more frequently recovered from locations near the toilet seat before cleaning when compared to E. coli and P. aeruginosa (P<0.05)(Table 6).There was a slight reduction in the cumulative number of bacteria on the surfaces observed 1 h after disinfection.Subsequent screening of the corridors after cleaning did not reveal the presence of E. coli, K. pneumoniae or A. baumannii, but S. aureus was still detected.

DISCUSSION
Despite a significant increase in antimicrobial resistance (AMR) and the spread of ARB such as E. coli, S. aureus, K. pneumoniae, A. baumannii and P. aeruginosa in hospital environments in low-to middle-income countries, as reported by WHO and Africa CDC [13,14], there is limited information available regarding the types of ARB and their associated ARGs colonizing major components of MSW, sludge, sewage and shared fomites in Nigeria.Since sewage and wastewater have been shown to accurately reflect the population's gut microbiota composition and antimicrobial use, it is crucial to conduct a comprehensive analysis of the bacterial communities present in community WWDs and MSW simultaneously.
All the samples from MSW, sewage, sludge and shared fomites that were screened yielded positive cultures, with a substantial proportion found in sludge and sewage.This finding is not surprising, as both sludge and sewage are rich in organic compounds that bacteria require for energy production.Sewage, which often contains faecal matter, harbours a slightly higher number of bacteria compared to sludge, which consists mainly of solids separated from wastewater originating from gutters, kitchens, toilets and nearby industries.Components of MSW, such as diapers and polythene bags used for sachet water, indiscriminately discarded in refuse dump sites, were found to harbour E. coli, K. pneumoniae and S. aureus at varying rates.Interestingly, none of the samples was contaminated with a single type of bacteria, suggesting a diverse environmental niche across the samples.While diapers are heavier and less likely to be blown away by the wind, polythene bags containing various bacteria could easily be transported through the air to other locations, including hospitals and school environments.Previous studies have also identified plastic debris [15], washroom floors [16], restroom surfaces [17], sewage soil [18], kitchen utensils [19], external surfaces, and alimentary tract of houseflies living on sewage [20] and food wastes [21] as surfaces and environmental reservoirs of bacterial pathogens such as S. aureus, E. coli, Enterococcus faecalis, K. pneumoniae and A. baumannii.Additionally, Butin et al. [22] isolated Staphylococcus capitis from several environmental sites inside the Neonatal Intensive Care Unit of a hospital.
The isolation of K. pneumoniae from all the samples is worrisome and indicates that this bacterium can readily establish itself in various environmental samples, similar to its persistence in clinical settings.Similarly, the recovery of E. coli and K. pneumoniae from nearly all components of MSW may suggest either easy transmission of these bacteria among MSW components or high faecal contamination in dump sites from which each component of MSW is collected.The frequent recovery of A. baumannii and K. pneumoniae from restroom floors and other fomites may be attributed to their versatile genetic machinery, which allows them to tolerate harsh environments, form biofilms and adhere to surfaces.Overall, the high volume of waste and dirt accumulated on restroom floors, shared by many students and ranging from tissue papers used for cleaning to sputum and urine discharged on the floors, may be a contributory factor to the persistent isolation of gastrointestinal bacteria [23].
The high resistance of bacteria isolated from all environmental wastes and shared fomites to penicillin is consistent with the high resistance often observed among clinical and veterinary isolates [24,25].Some bacteria (P.aeruginosa, A. baumannii and S. aureus) isolated from sewage and sludge have also developed resistance to fluoroquinolones, tetracyclines and colistin.Similar observations in animals have been reported by Egbule and Yusuf [5], where isolates from poultry and cattle dung in environments have developed significant resistance to common antibiotics.The majority of bacteria isolated are susceptible to other antibiotics, despite exhibiting some level of resistance to the same antibiotics among clinical samples.In line with this finding, resistance to penicillin and fluoroquinolones by bacteria isolated from landfills and laboratories has been reported to have increased in studies Table 6.Detection of bacteria on restroom floors and corridors for four consecutive days before and 1 h after cleaning and disinfection process conducted in Ghana and Zimbabwe [26,27].Kanamycin, gentamicin and ampicillin resistance by A. baumannii have been widely reported among clinical isolates [28] as well as intrinsic resistance to chloramphenicol [29].
The production of ESBL by E. coli and K. pneumoniae isolated from some components of MSW, WWDs and shared fomites aligns with the high-level recovery of ESBL-E.coli from clinical samples in the same environment [9,30].This could indicate the presence of hospital waste in the environments sampled in this study.However, the detection of ESBL-K.pneumoniae in sewage and ESBL-P.aeruginosa in diapers and food waste could suggest contamination from households.
The presence of bla NDM-1 and bla KPC in two different K. pneumoniae isolates and bla IMP in one non-ESBL E. coli isolated from sewage and sludge, respectively, is of public health concern and corresponds to the findings of previous studies in different regions of Nigeria where bla NDM and bla KPC have been reported in clinical samples [31,32].It is worth noting that A. baumannii genotypically harbouring bla OXA-51-like and bla CMY-2 genes is phenotypically susceptible to imipenem and cephalosporins, probably due to the lack of insertion sequence ISAba1 adjacent to bla OXA-51-like , which serves as a promoter for carbapenem resistance expression.
Although the source of these bacteria is not clear at present, it could be linked to close interactions between humans, animals and waste, facilitating the exchange of pathogens through the activities of waste scavengers, open defecators in waste disposal sites, and runoff from disposal sites to houses and vice versa (see Fig. 2).Previous reports have indicated the presence of resistant genes in river water in Vietnam [33], seepage and tap water in India [34], and bla IMP-1 in Tunisian rivers [35].Another possible reason is that freely moving wild animals have access to waste disposal sites, and ARB shed by these animals may contaminate the soil and other waste dumped on it.Similarly, wild animals can come into close contact with sewage and sludge, leading to the widespread dissemination of ARB and ARGs originally found in hospitals to the environment [36].
Finally, the partial failure of the disinfection process to completely remove bacteria, specifically ARB, may be explained by a decreased susceptibility of the bacteria to the disinfectants used during cleaning.Shorter contact times, inappropriate concentrations and technical difficulties in reaching hidden areas of the restroom may have limited the effectiveness of the disinfectants in exerting their bactericidal effects, thus favouring the persistence of bacteria.

CONCLUSION
The present study has established that municipal solid waste indiscriminately discharged in some urban cities in Nigeria, as well as sewage, sludge and frequently shared fomites, often harbour ARB such as S. aureus, E. coli, K. pneumoniae, P. aeruginosa and A. baumannii.These bacteria display resistance to penicillin, tetracycline and fluoroquinolones.Furthermore, ESBL-producing strains of K. pneumoniae, E. coli, P. aeruginosa and A. baumannii, as well as other non-ESBL producers, carry one or more ARGs, including bla KPC , bla NDM-1 , bla CMY-2 and bla IMP , which are commonly found in hospital-acquired strains.Notably, the disinfection process employed in the university did not effectively reduce the levels of ARB.Therefore, we advocate for an informed One Health strategy to address the role of the environment in the emergence and spread of AMR in Nigerian urban communities.

Funding Information
The author(s) received no specific grant from any funding agency.
The Microbiology Society is a membership charity and not-for-profit publisher.
Your submissions to our titles support the community -ensuring that we continue to provide events, grants and professional development for microbiologists at all career stages.The WGS of the A. baumanniiwere carried out.The assembled data were analyzed to identify the bacterial species and gather additional information.Bacterial species was determined using tools Kraken, and BLAST against reference databases In addition, please ensure that the sequenced ARGs sequences (Line 153) are also deposited with the accession provided.
The sequenced ARGs were deposited through BankIt of NCBI.We hoped to get Accession number next week Reviewer 1 How the identification of the bacterial isolates to the species level was made?API kits were used.It has been mentioned in the manuscript Data analysis by using SPSS mentioned in the methods are not represented in the results..

Done
Did the Authors design the primers ?If not so they have to add the reference to table 1.
References added, except for the one designed by the authors the cycle conditions of the PCR are better to be added to the references.

Done
In table 1 the names of the primers should be written in italics.

Done
In the results: It is not clear how was the % beside each number calculated, so it is better add the total number (100%) in the last column or the last row Erroneously done, but has been corrected Although the study design seems sound, the type of the study and the sample size issue has to be clarified.
The selection was based on pilot studies conducted in which All the 5 bacteria and Enterobacter, Citrobacter, Proteus were targeted, but the selected had the highest prevalence It seems to be more interesting if screening was done for ESKAPE pathogens and it will be a more contextualized methods.

Noted
There are insufficient details of the used methods e.g.laboratory procedures followed are not clarified specially: methods for isolation and identification of selected bacteria ; antibiotic susceptibility testing.
This has been rectified * The work is fundamentally sound.But there may be a bias in some results regarding Efficacy of floor disinfection process routinely used in reducing burden of selected bacteria as there are many ignored factors as type of disinfectant used, concentration and contact time.
The same disinfectants were used and the same procedure.Our target is to know if the routine procedure used is efficient enough to remove ARBs.
Another study (a year later) surveyed different brands of disinfectants, and different concentrations were used.The data was used to adjust the practice I strongly advise the author to rewrite the methodology section to produce a more contextualized and scientific manuscript.

This has been done
The English needs attention throughout the manuscript.
We have improved the English Comments: This is a study that would be of interest to the field and community.The reviewers have highlighted major concerns with the work presented.Please ensure that you address their comments.Please include more rigour criteria and resources in your methods section, as highlighted by the SciScore reports.Including RRIDs and negative statements to explain why things were not performed should increase the rigour and reproducibility of your work.You can find tips on how to improve your article here: https:// sciscore.com/ reports/ Core-Report.php Please provide more detail in the Methods section and ensure that software is consistently cited and its version and parameters included.
1. the cycle conditions of the PCR are better to be added to the references.In table 1 the names of the primers should be written in italics.In the results: It is not clear how was the % beside each number calculated, so it is better add the total number (100%) in the last column or the last row

Fig. 1 .
Fig. 1.Locations where samples of MSW and waste water were collected.The sampling points were situated near residential areas to which humans (passersby and scavengers) and animals have free access.Each component of MSW was picked from wastes deposited for more than 1 week.Yellow arrows indicate sites where waste water derivatives were sampled (sewage and sludge).
All values are expressed as a percentage rounded up to the nearest whole number.nt=not tested, WWD=waste water derivatives, MSW=municipal solid waste, SF=shared fomites.Breakpoints of Comité de l'Antibiogramme de la Société Française de Microbiologie recommendations were used for tigecycline, gentamicin and tetracycline for A. baumannii.

Fig. 2 .
Fig. 2. The big picture of a how municipal solid wastes and waste waters indiscriminately discharged into the environment in Nigeria generate and transmit ARB and ARGs.The diagram explains how different human activities are being carried out in the communities (such as burning wastes, scavenging by humans and animals).Airborne bacterial pathogens are released during burning, wind blowing, and loading and unloading of garbage by waste dischargers, and scavenging humans and animals and could lead to the spread of ARB to the nearby human population and fomites.

VERSION 1 Editor
recommendation and comments https://doi.org/10.1099/acmi.0.000641.v1.5 © 2023 Beedessee G.This is an open access peer review report distributed under the terms of the Creative Commons Attribution License.Girish Beedessee; University of Cambridge, UNITED KINGDOM Date report received: 03 August 2023 Recommendation: Major Revision

Table 1 .
Genes amplified in resistant Gram-negative bacteria isolated from environmental wastes and fomites

Table 2 .
Overall recovery of bacteria from MSW, WWD and SF

Table 3 .
Percentage susceptibility of bacteria isolated from different MSW, waste water derivatives and shared fomites to multiple antibiotics

Table 5 .
Detection of ARGs in randomly selected isolates from different sources na=not applicable; ARGs=antibiotic resistance genes.

Table 4 .
Screening of bacteria isolated from MSW, WWD and SF for MRSA and ESBL production

Table 3 :
All abbreviations mentioned in the table should be written just below the table so the reader won't go back to the text to find it.

Table 3 :
All abbreviations mentioned in the table should be written just below the table so the reader won't go back to the text to find it.Minor comments: Line 33: methicillin resistant not resistance.line76:referencesaretobemoved to the end of the sentence.Please rate the manuscript for methodological rigour GoodPlease rate the quality of the presentation and structure of the manuscript GoodTo what extent are the conclusions supported by the data?Strongly supportDo you have any concerns of possible image manipulation, plagiarism or any other unethical practices?NoIs there a potential financial or other conflict of interest between yourself and the author(s)?NoIf this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?Yes